Acyloxymethyl esters of alpha-ureidocyclo-hexadie nylalkylene-cephalosporins

ABSTRACT

New acyloxymethyl esters of Alpha ureidocyclohexadienylalkylene-cephalosporins of the general formula   ARE EFFECTIVE AS ANTIBACTERIAL AGENTS AND SHOW PROLONGED ACTION AS WELL AS OTHER IMPROVED RESULTS.

United States Patent 1 [111 3,708,479

Welch et al. 1 Jan. 2, 1973 [s41 ACYLOXYMETHYL ESTERS 0F 3,352,851ll/l967 Fosker ..260/239.l ALPHA-UREIDOCYCLO-HEXADIE 3,250,679 5/l966Jansen el al ..260/239.l

NYLALKYLENE-CEPHALOSPORINS Primary Examiner-Nicholas S. RizzolflvenlOrSIAmold welch, Princemn. Attorney-FLawrence S. Levinson, MerleJ. Smith,

Ni 08540; Jmph Edward Donald .I.Perrella and Burton Rodney fini, NorthBrunswick, N..I.,

grjderlck F. Giarrusso, Belle Mead, 57 ABSTRACT [73] Assignecz Squibb &Sons Inc New New acyloxymethyl esters of n-ureidocyclohex- Yorkadienylalkylene-cephalosporins of the general formula [22] Filed: Oct.26, 1970 0 s ll 21 Appl. No.: 84,170 3 R NH O=J7--N\ c cH,x [52] U.S.Cl. ..260/243 C, 424/246 n 5 l] m. Cl. ..C07d 99/24 Nu, 0=Jv 0 Clh-0-:-n [58] Fleld of Search ..260/243 C are effecmm as antibaclcrial agentsand Show p longed action as well as nlher improved results, I56]References Clled UNITED STATES PATENTS 7 Clalms, No Drawings 3,485,8l912/1969 Weisenborn et al ..260/239.l

ACYLOXYMETI-IYL ESTERS OF ALPHA- UREIDOCYCLO-HEXADIE NYLALKYLENE-CEPHALOSPORINS SUMMARY OF THE INVENTION s cm)n-cni lNfi-cn-cfi cm H IJH0=)-lil (J-CHrX 3:0 C 1 111: O=0-CHr-Oi-R which are antibacterial agentseffective against 5 variety of bacterial pathogens, both gram-positiveand gram-negative bacteria, and which give rise to higher blood levelsof the antibiotic.

R in formula 1 represents hydrogen, lower alkyl or lower alkoxy. R islower alkyl or monocyclic carbocyclic arylor aralkyl (the aryl portionsof the last two radicals being monocyclic carbocyclic aryl and the alkylbeing lower alkyl of up to seven carbons).

X is hydrogen, lower alkoxy, lower alkanoyloxy, aroyloxy oraralkanoyloxy, (the aryl and aralkyl portions of the last two groupsbeing the same as defined for R, e.g., benzoyloxy, phenyl-loweralkanoyloxy like phenylacetoxy), the radical of a nitrogen base or aquaternary ammonium radical. n is 0, l, 2, 3 or 4.

Preferred members in each group include R as hydrogen; n as 0 or 1,especially 0; R as t-butyl or phenyl, especially t-butyl; X as hydrogenor lower alkanoyloxy, especially acetoxy.

DETAILED DESCRIPTION OF THE INVENTION The symbol R in formula Irepresents a lower alkyl group, e.g., a branched or straight chainsaturated aliphatic hydrocarbon group having up to seven carbons in thechain, including methyl, ethyl, propyl, isopropyl, butyl, isobutyl,t-butyl, amyl and the like and also lower alkoxy groups wherein similaralkyl groups are attached to an oxygen.

R completes the ester group and represents a lower alkyl group of thetype defined previously, preferably t butyl. R may also representmonocyclic carbocyclic aryl or aralkyl groups like phenyl, benzyl,phenethyl, phenylisopropyl or the like. Thus acyloxymethyl ester likeacetoxymethyl, propionyloxymethyl, isopropionyloxymethyl,butyryloxymethyl, pivaloyloxymethyl, valeroyloxymethyl,benzoyloxymethyl,

phenacetyloxymethyl esters and the like are included.

The symbol n represents 0, l, 2, 3 or 4, preferably the first two. X ishydrogen, lower alkoxy, e.g., methoxy, ethoxy, propoxy, isopropoxy orthe like, lower alkanoyloxy, e.g., acetoxy, propionyloxy or the like orthe radical of an amine, e.g., N-pyridinium, or the like.

The new compounds of this invention are preferably produced from eithera 7-aminocephalosporanic acid compound of formula ll or salt thereof orfrom a 7-[2-ureido2-(1,4-cyclohexadienylalkyl)acetamidol-cephalosporanic acid of formula illor salt thereof.

R, X and n have the meanings already defined.

The salts are those formed with the carboxyl group and are known e.'g.,alkali metal salts, alkaline earth metal salts, salts with organicamines etc.

The acyloxymethyl ester of either a compound of formula II or formulall] is produced, e.g., by a method adapted from that of Daehne et al.,Jour. Med. Chem. [3, 607 I970), from either type of compound by reactionwith a halomethyl ester halCH OCoR wherein R has the meaning alreadydefined and hal is halogen, preferably chlorine or bromine.

The acid or salt of either formula ll or formula III is treated with thehalomethyl ester in a molar ratio of about lzl to 1:2 in an inertorganic solvent like dimethylformamide, acetone, dioxane, benzene or thelike at about ambient temperature or below.

The products resulting from the reaction of a compound of formula IIIwith a halomethyl ester of formula IV may beisolated as slightly solublesalts with inorganic or organic acids such as hydrochloric acid orptoluenesulfonic acid.

In the case ofa compound of formula II reaction with an acyl halomethylester provides a corresponding acyloxymethyl ester. The resulting esteris then reacted with a I,4-cyclohexadienyl-a-aminoalkanoic acid, thecarbamyl derivative or an N-protected derivative thereof as described inmore detail below.

The starting compounds of formula Ill are produced as described in thecopending application of Raymond Curry Erickson Ser. No. 877,488, filedNov. 17, 1969, by first forming a compound of the formula and thentreating the compound of formula V with a cyanate, e.g., an alkali metalcyanate such as potassium cyanate or with carbamyl phosphate.

The substances of formula V are produced as described in US. Pat. No.3,485,819, issued Dec. 23, I960, from a 7-aminocephalosporanic acidcompound of the formula wherein R and X have the meanings alreadydescribed. The latter are obtained as described in U.S. Pat. No.

The carbamylation is effected by dissolving or suspending the a-aminoacid compound in aqueous medium and the cyanate or carbamyl phosphate isslowly added. Heat, e.g., up to about 80 C., may be used to acceleratethe reaction. The pH of the reaction mixture is preferably kept on theacid side, e.g., within the range of about to 6.9. The product mayusually be precipitated by acidification and chilling.

As indicated, the carbamylation reaction may be carried out as the laststep of the synthesis, as preferred, or at any earlier stage of thesynthesis.

It will be appreciated that certain of the compounds of this inventionexist in various states of solvation as well as in different opticallyactive forms. The various forms as well as their mixtures are within thescope of this invention.

Ordinarily the new compounds of this invention derived from D-a-aminoacids or derivatives thereof are more active than the correspondingcompound derived from the L-form or DL-form. The configuration of thea-carbon of the a-amino acid used in the synthesis is retained in theproduct.

The compounds of this invention have a broad spectrum of antibacterialactivity against both gram positive and gram negative organisms such asStaphylococcus aureus, Salmonella schottmuelleri, Proteus vulgaris,Escherichia coli and Streptococcus pyogenes. They may be used asantibacterial agents in a prophylactic manner, e.g., in cleaning ordisinfecting compositions, or otherwise to combat infections due toorganisms such as those named above, and in general may be utilized in amanner similar to chephalosporin C, cephalothin and othercephalosporins. For example, a compound of formula I may be used invarious animal species in an amount of about 1 to 200 mgJkg. daily,orally or parenterally, in single or two to four divided doses to treatinfections of bacterial origin. By way of illustration the PD,subcutaneously in mice in a single administration is of the order of 5mgJkg. against Streptococcus. The oral form gives a prompt high bloodlevel which is maintained for relatively long periods.

Up to about 600 mg. of a compound of formula I or a salt thereof may beincorporated in an oral dosage form such as tablets, capsules or elixirsor in an injectable form in a sterile aqueous vehicle prepared accordingto conventional pharmaceutical practice.

in cleaning or disinfecting compositions, e.g., in barns or dairyequipment, a concentration of about 0.0l to 1 percent by weight of suchcompounds admixed with, suspended or dissolved in conventional inert dryor aqueous carriers for application by washing or spraying may be used.

They are also useful as nutritional supplements in animal feeds.

The following examples are illustrative. All temperatures are on thecentigrade scale.

EXAMPLE I a. D-2-Amino-2-( l ,4-cyclohexadien-l-yl)acetic acid Asolution of 11.0 g. (72.7 mmol.) of D-phenylglycine in 900 ml. distilledammonia (which has been treated with 45 mg. lithium after distillationto destroy traces of moisture) is slowly diluted with 370 ml. dry I-butyl alcohol.

Over a period of 2 hours, L65 g. lithium (3.27 eq.) is added in smallportions until a permanent blue color is obtained. The blue reactionmixture is then treated with 38 g. of triethylamine hydrochloride. Theammonia is allowed to evaporate at room temperature overnight and theresidual solvent is evaporated at reduced pressure. The white residue istaken up in a small amount of methanol-water and added to 4 t. of cold1:] chloroform acetone to precipitate the crude product. After 20minutes stirring the suspension is filtered and the white filter cakedried in vacuo; the filter cake is then pulverized and submitted oncemore to the precipitation process from 1:1 chloroform-acetone. D-2-amino-2-(l,4-cyclo-hexadien-l-yl)acetic acid is obtained as a whitecrystalline product, m.p. 297 (dec.).l b. Methyl acetoacetic esterenamine of N-2-amino-2- (l,4-cyclohexadien-l-yl)acetic acid sodium salt306 mg. D-2-amino-2-(1,4-cyclohexadien-lyl)acetic acid (2.00 mmols.) aredissolved by warming in a solution of I08 mg. of NaOCH, (2.00 mmols.) in4.3 ml. reagent grade MeOH. 255 mg. (0.24 ml. 2.20 mmols.) methylacetoacetate are added and the mixture refluxed for 45 minutes. The MeOHis almost totally stripped off in vacuo. 5 ml. benzene are added anddistilled off to a small residual volume. The addition and distillationof benzene is repeated to insure complete removal of the MeOH and water.The product crystallizes our overnight from a small residual volume ofbenzene. It is filtered off, washed with benzene, and dried in vacuo.7-[D-2-Amino-2-(1,4-cyclohexadien-lyl)acetamido]cephalosporanic acid 452mg. of 7-aminocephalosporanic acid (7ACA) are stirred well in 2.5 ml. ofwater while 0.23 ml. triethylamine are gradually added with the pH keptunder 8.0. Final pH is 7.4; 0.85 ml. acetone are added and the solutionis kept at l 0 C.

469 mg. methyl acetoacetate enamine of D-Z-amino-2-(l,4-cyclohexadien-l-yl)acetic acid sodium salt, are stirred in 4.25ml. acetone at 20 C. A microdrop of N-methylmorpholine is added followedby the slow addition of 198 mg. of ice cold ethyl chloroformate. 0.43ml. of water is added at this point. The reaction mixture is stirred for10 minutes at 20 C.

The solution of mixed anhydride is then added to the 7-ACA solution. Thesolution is stirred for 30 minutes at l0 C., then raised to roomtemperature, acidified to pH 2.0 with diluted HCl and, with goodstirring the pH is kept at that level for 10 minutes.

The solution is then extracted with 5 ml. xylene. The aqueous layer islayered with 5 ml methyl isobutyl ketone and the pH adjusted to 5.0 withl N NaOH and chilled overnight. The resulting crystals of 7-[D-2-amino-2-( l ,4-cyclohexadien-1-yl)- acetamidolcephalosporanic acid arewashed with water and air dried.

d. 7-[D-2-ureido-2-(1,4-cyclohexadien-lyl)acetamidolcephalosporanic acidand potassium salt To a suspension of mg. of the product of part c in7.5 ml. of water there are added mg. of potassium cyanate with stirring.The reaction mixture, which soon clears, is incubated at 22-24 and thepH maintained filtered off,

between 5.0 and 6.9 by frequent dropwise additions of 1.0 N hydrochloricacid with stirring. After 5 hours, the solution is adjusted to 10.0 ml.,acidified to pH LB with 1.0 N. hydrochloric acid, and extractedsuccessively with four 75 ml. portions of ethyl acetate. The ethylacetate is washed four times with ml. portions of water adjusted to pH2.0 with hydrochloric acid, filtered and evaporated at l0-20C. in vacuoto give 7- [D-2-ureido-2-( 1,4-cyclohexadienlyl)acetamido]cephalosp0ranic acid as an amorphous white powder.

The potassium salt is obtained by suspending the powder in 80 ml. ofwater and treated with one equivalent of 0.1 N aqueous potassiumhydroxide solution which is added with vigorous stirring. The solutionis evaporated to dryness in vacuo at 25-30 to obtain the potassium salt.

e. Pivaloyloxymethyl ester of 7-[D-2-ureido-2-(l,4- cyclohexadienl -yl)acetamidolcephalosporanic acid 3.0 gm. mmol.) of chloromethylpivalateare combined with 3.90 gm. (l0 mmol.) of the potassium salt of part d,0.4 ml. of a five percent aqueous sodium iodide and I70 ml. of acetone.The whole is stirred for l6 hours under an argon atmosphere followed byone hour of heating at reflux temperature. The mixture is cooled,filtered and concentrated at reduced pressure. The residue ispartitioned between 250 ml. ethyl acetate and 100 ml. of dilute aqueoussodium bicarbonate solution. The organic layer is washed with wateruntil the washes are at pH7. After drying over sodium sulfate, theorganic solution is concentrated at reduced pressure. The oily residueis scrubbed with ether to produce the solid pivaloyloxymethyl ester of7-[D-2- ureido-2-( l,4-cyclohexadienl yl)acetamido]cephalosporanic acid.

EXAMPLE 2 EXAMPLE 3 Pivaloyloxymethyl ester of 7-[D-2-ureido-2-(L4-cyclohexadienl -yl )acetamido]cephalosporanic acid 300 mg. (2 mmol.) ofchloromethylpivalate areadded to a mixture of 390 mg. (1 mmol.) of theproduct of Ex ample l a (potassium salt) and l5 ml. ofdimethylformamide. The whole is stirred under an argon atmosphere for 24hours. 150 ml. of ethyl acetate are added and the whole is filtered. Thefiltrate is washed with three portions of 5 percent aqueous sodiumbicarbonate solution and with water until the water is at pH 7. Theproduct is then dried over sodium sulfate and concentrated at reducedpressure. The residual product, the pivaloyloxymethyl ester of7-[2-ureido-2- l ,4-cyclohexadienl -yl )acetamidolcephalosporanic acid,is scrubbed with hexane and dried.

EXAMPLE 4 By substituting 20 mmol. of chloromethylbenzoate for thechloromethylpivalate in the procedure of Example l e, thebenzoyloxymethyl ester of 7-[D-2-ureido-2- l ,4-cyclohexadienl -yl)acetamidolcephalosporanic acid is obtained.

EXAMPLE 5 a. D-2-amino-3-( l ,4-cyclohexadienl -yl)propionic acid 12.0g. of D-phenylalanine are substituted for the D- phenylglycine in theprocedure of Example I to obtain D-2-amino-3-( l ,4-cyclohexadienl-yl)propionic acid as a white powder.

b. Methyl acetoacetate ester enamine of D-2-amino-3-(l,4-cyclohexadien-l-yl)propionic acid sodium salt This product isobtained by substituting 330 mg. of the product of part a above in theprocedure of Example l b.

c. 7-[ D-2-amino-3-( l ,4-cyclohexadienlyl)propionamido]-cephalosporanic acid 493 mg. of methyl acetoacetateenamine of D-2- amino-3-( l ,4-cyclohexadienl -yl)propionic acid sodiumsalt (1.715 mmol.) are substituted for the methyl acetoacetate enamineof D-2amino-2-(1,4-cyclohexadien-l-yl)acetic acid sodium salt in theprocedure of Example I c.

The mixed anhydride is added to the 7-ACA, stirred for 30 minutes at -10C., brought to room temperature, acidified to pH 2.0 as in Example 1 c.The solution is then extracted with 5ml. of xylene. The aqueous layer islayered with 5 ml. of methyl isobutyl ketone and the pH is adjusted to5.0 with l N NaOH. The aqueous layer is then lyophilized to give7-[D-2-amino- 3-( l ,4-cyclohexadienl -y|)propionamido1cephalosporanicacid.

d. 7-[D-2-ureido-3-( l,4-cyclohexadienl yl)propionamidol-cephalosporanicacid and salt By utilizing the product of part c in the procedure ofExample I d, 7-[D-2-ureido-3-(,4-cyclohexadienlyl)-propionamido]cephalosporanic acid and the potas'sium salt are obtained.

e. Pivaloyloxymethyl ester of 7-[D-2-ureido-3-( l ,4-cyclohexadien-l-yl)propionamidolcephalosporanic acid By utilizing thepotassium salt of part d above in the procedure of Example 1 e, thepivaloyloxymethyl ester of 7-[D-2-ureido 3-( l ,4-cyclohexadienlyl)propionamidolcephalosporanic acid is obtained.

EXAMPLE 6 Acetoxymethyl ester of 7-[D-2-ureido-2-(4-methoxy'l.4-cyclohexadienl -yl )propionamidolcephalo sporanic acid Bysubstituting l4.2 g. (72.7 mmol.) of D-O-methyltyrosine for thephenylglycine in the procedure of Example l a, D-2-amino-3-(4-methoxy-l,4-cyclohexadienl -yl)propionic acid is obtained as a white crystallineproduct, m.p. 227 C.

Then by using this compound in the procedure of Example l, parts b, cand d, and by substituting 20 mmol. of chlo romethyl-acetate for thechloromethyl pivalate in part e, the above named product is obtained.

EXAMPLE 7 Pivaloyloxymethyl ester of 7-[D-2-ureido-3-(l,4-cyclohexadien- 1 -yl )-propionamido]3-desacetoxycephalosporanic acid Byutilizing the procedure of Example 5, but substituting 356 mg. of3-desacetoxy-7- aminocephalosporanic acid for the 7-ACA, the aboveproduce is obtained.

EXAMPLE 8 Pivaloyloxymethyl ester of 7-[DL-2-ureido-2-(4- methyl-l,Lcyclohexadien- 1 -yl )acetamido lcephalospo ranic acid By substitutingDL-4-methylphenylglycine for the phenyl-glycine in part a and otherwisefollowing the procedure of Example 1, the above product is obtained.

EXAMPLE 9 Pivaloyloxymethyl ester of 7-[DL-2-ureido-2-(4- methyl-l,4-cyclohexadien-1-yl)-3-desacetoxycephalosporanic acid By substitutingDL-4-methylphenylglycine for the phenyl-glycine in part a andsubstituting 7-ADCA for 7-ACA in part c, then otherwise following theprocedure of Example 1, the above named product is obtained.

EXAMPLE 10 a. By substituting 10 mmol. of 7-ACA and adding 20 mmol. oftriethylamine but otherwise following the procedure of Example 1 e thepivaloyloxymethyl ester of 7-ACA is obtained.

b. 647 mg. of 7-aminocephalosporanic acid pivaloyloxymethyl ester arestirred well in 3.0 ml. of acetone and the solution is kept at 1 C.

469 mg. of the methyl acetoacetate enamine of D-2- amino-2-( l,4-cyclohexadienl -yl)acetic acid sodium salt, are stirred in 4.25 ml.acetone at 20 C. A microdrop of N-methyl-morphoiine is added followed bythe slow addition of 198 mg. of ice cold ethyl chloroformate. 0.43 ml.of water is added at this point. The reaction mixture is stirred for 10minutes at 20 C.

The solution of mixed anhydride is then added to the 7-ACA estersolution. The solution is stirred for 30 minutes at 10 C., then raisedto room temperature, acidified to pH 2.0 with diluted HCI and, with goodstirring the pH is kept at that level for 10 minutes.

The solution is then diluted with ml. water and extracted with 5 ml.xylene. The aqueous layer is layered with ml. ether and the pH adjustedto 7.0 with l N NaOH. The ether layer is dried over sodium sulfate andthen evaporated at reduced pressure to deposit the product, thepivaloyloxy-methyl ester of 7-[D-2-amino- 2-( l ,4-cyclohexadienl -yl)-acetamidolcephalosporanic acid.

c. To a suspension of 85 mg. of the product of part b in 10 ml. ofacetone:water (1:1) there are added 150 mg. of potassium cyanate withstirring. The reaction mixture is incubated at 2224 and the pHmaintained between 5.0 and 6.9 by frequent dropwise additions of 1.0hydrochloric acid with stirring. After 5 hours, the

solution is adjusted to 10.0 ml., acidified to pH 1.8 with 1.0 Nhydrochloric acid, and extracted successively with four ml. portions ofethyl acetate. The ethyl acetate is washed four times with 10 ml.portions of water adjusted to pH 2.0 with hydrochloric acid, fil teredand evaporated at l0-20 C. in vacuo to give 7-[D-2-ureido-2-(1,4-cyclohexadien-lyl)acetamido]cephalosporanic acidpivaloyloxymethyl ester as an amorphous white powder.

EXAMPLE 1 l By substituting 10 mmol. of 7-ADCA for the 7-ACA in theprocedure of Example 10 a, the same product is obtained as in Example 2.

EXAMPLE 12 a. A suspension of 1.0 g. of DL-2-amino-3-( 1,4-cyclohexadien-l-yl)-acetic acid in 10.0 ml. of water is treated with0.59 g. of potassium cyanate and heated at C. with stirring until aclear solution develops. The solution is cooled, incubated at 24 C. for1 8 hours and then acidified with 2.0 N hydrochloric acid to precipitateDL-2-ureido-3-( 1,4-cyclohexadienl yl)acetic acid which is washed withcold water and dried over calcium chloride in vacuo.

b. 647 mg. of 7-aminocephalosporanic acid pivaloyloxymethyl ester arestirred well in 3.0 ml. of acetone and the solution is kept at 10 C.

349 mg. of DL-2-ureido-2-(1,4-cyclohexadien-1- yl)acetic acid arestirred in 4.25 ml. acetone at 20 C. A microdrop of N-rnethylmorpholineis added followed by the slow addition of 198 mg. of ice cold ethylchloroformate. 0.43 ml. of water is added at this point. The reactionmixture is stirred for 10 minutes at 20 C.

The solution of mixed anhydride is then added to the 7 ACA estersolution. The solution is stirred for 30 minutes at 10 C., then raisedto room temperature, acidified to pH 2.0 with diluted HCl and, with goodstirring the pH is kept at that level for 10 minutes. The solution isdiluted with 10 ml. water and extracted with 20 ml. ethyl acetate. Theorganic layer is washed with cold 1 percent aqueous hydrochloric acidand cold 1 percent sodium bicarbonate. After drying over sodium sulfate,evaporation at reduced pressure the product, the pivaloyloxy-methylester of 7-[DL-2-ureido.-2-( 1 ,4- cyclohexadienl -yl)-acetamidolcephalosporanic acid is deposited.

EXAMPLE l3 Benzoyloxymethyl ester of 7-[D-2-Amino-2-(l,4- cyclohexadienlyl)acetamido]cephalosporanic acid a. 33 mg. (2 mmol.) ofchloromethylbenzoate are added to a mixture of 35 mg. 1 mmol.) of theproduct of Example 1 c and 15 ml. of dimethylformamide. The whole isstirred under an argon atmosphere for 24 hours. ml. of ethyl acetate areadded and the whole is filtered. The filtrate is washed with threeportions of 5 percent aqueous sodium bicarbonate solution and with wateruntil the water is at pH 7. The product is then dried over sodiumsulfate and concentrated at reduced pressure. The residual product,-thebenzoyloxymethyl ester of 7-D-[2-amino-2-( l ,4cyclohexadien lyl)acetamido]cephalosporanic acid, is scrubbed with hexane and dried.

9 b. Benzoyloxymethy] ester of 7-[D-2-ureido-2-(li4- cyclohexadienl -y])acetamido]cephalosporanic acid The product of part a is treated as inExample 1 d to obtain the above named product.

EXAMPLE l4 EXAMPLE IS a. D-2-Amir1o-3-( l,4-cyclohexadien-l acid 12.0 g.of D-phenylalanine are substituted for the D- phenylglycine in theprocedure of Example I a to obtain D-2-amino-3-( l ,4-cyclohexadienl-yl)propionic acid as a white powder. b. Methyl acetoacetate esterenamine of D-2-amino-3- l,4-cyclohexadienyl)propionic acid sodium saltThis product is obtained by substituting 330 mg. of the product of parta above in the procedure of Example l b.

c. 7-[D-2-amino-3-( l,4-cyclohexadien-lyl)propionamidol-cephalosporanicacid 493 mg. of methyl acetoacetate enamine of D-2-amino-3-(l,4-cyclohexadienyl)propionic acid sodium salt (L715 mmol.) aresubstituted for the methyl acetoacetate enamine ofD-2-amino-2-(l,4-cyclohexadien-l-yl)acetic acid sodium salt in theprocedure of Example 1 c.

The mixed anhydride is added to the 7-ACA, stirred for 30 minutes at C.,brought to room temperature, acidified to pH 2.0 as in part c. Thesolution is then extracted with 5 ml. of xylene. The aqueous layer islayered with 5 ml. of methyl isobutyl ketone and the pH is adjusted to5.0 with l N NaOl-l. The aqueous layer is then lyophilized to give7-[D-2-ami'no-3-(l,4- cyclohexadien-l-yl)propionamido]cephalosporanic-yl )propionic acid. d. 7-[ D-2-ureido-3-( l ,4-cyclohexadienlyl)propionido]cephalosporanic acid By utilizing the product of part cabove in the procedure of Example I d, the pivaloyloxymethyl ester of7-[D-2(-amino-3-( 1,4-cyclohexadien-l yl)propionamido]-cephalo-sporanicacid is obtained.

e. Pivaloyloxymethyl ester of 7-[D-2-ureido-3-(l,4- cyclohexadienl-yl)propionamido ]cephalosporanic acid By utilizing the product of part0 above in the procedure of Example I e, the above named product isobtained.

EXAMPLE l6 Acetoxymethyl ester of7-[D-2-ureido-2-(4-methoxyl,4-cyclohexadienl -yl )propionamidolcephalosporanic acid potassium salt By substituting 14.2 g. (72.7 mmol.) ofD-O-methyltyrosine for the phenylglycine in the procedure of Ex ample la, D-2-amino-3-(4-methoxy-l,4-cyclohexadienylJpropionic acid is obtainedas a white crystalline product, m.p. 227 C.

Then by using this compound in the procedure of Example parts b, c andd, and by substituting l.l gm. of chloromethyl-acetate for thechloromethyl pivalate in part e, the above named product is obtained.

EXAMPLE [7 Pivaloyloxymethyl ester of 7-[D-2-ureido-3-( 1,4-cyclohexadienl -yl )propionamido]-3-desacetoxycephalosporanic acid Byutilizing the procedure of Example 15, but substituting 356 mg. of3-desacetoxy-7- aminocephalosporanic acid for the 7-ACA, the aboveproduct is obtained.

EXAMPLE l8 Pivaloyloxymethyl ester of 7-[DL-2-ureido-2-(4- methyll,4-cyclohexadienl -yl)acetamido]cephalospo ranic acid By substitutingDL-4-methylphenylglycine for the phenyl-glycine in part a and otherwisefollowing the procedure of Example I, the above product is obtained.

EXAMPLE l9 Pivaloyloxymethyl ester of 7-[DL-2-ureido-2-(4- methyl- I,4-cyclohexadien-1-yl)-3-desacetoxycephalosporanic acid By substitutingDL-4-methylphenylglycine for the phenyl-glycine in part a andsubstituting 7-ADCA for 7-ACA in part c, then otherwise following theprocedure of Example 1, the above product is obtained.

EXAMPLE 20 Phenylacetoxymethyl ester of 7-[DL2-ureido-2-(4- methoxy- I,4-cyclohexadienl yl)acetamido ]cephalosporanic acid By utilizingphenylacetoxymethyl chloride in part e and DL-(4-methoxy)phenylglycinein part a of Example l, the above named compound is obtained.

EXAMPLEZI 7-[ D-2ureido( l ,4-cyclohexadienl-yl)acetamidolcephalosporanic acid pivaloyloxymethyl ester, sterile 250 mg.Lecithin powder, sterile 50 gm. Sodium carboxymethylcellulose, sterile20 gm,

The sterile powders are aseptically blended, subdivided, filled intosterile vials and sealed. The addition of 1 ml. of water for injectionto the vial provides a suspension for intramuscular injection.

EXAMPLE 22 The following ingredients are admixed:

7-[ D-Z-ureido-2-( l ,4yclohexadien' l -yl )acetamido lcephalosporanicacid pivaloyloxymethyl ester 250 gm. Lactose 56.9 gm. Magnesium stearale3. l gm.

The mixed ingredients are subdivided and filled into 1 l 12 L000 number2 gelatin capsules each containing a total 1. A compound of the formulaof 310 mg. with 250 mg. of active substance.

EXAMPLE 23 11, c Hi JNII-CH(IJH ()IIE Tablets are prepared from thefollowing ingredients: R 11;}; O=d1-N C-CIIX l s I 7-[ D-2-ureido-( l,4-cyclohexadien- NH? O i-O.OH2O R l-yl)acetarnidolcephalosporanic acidpivaloyloxymethyl ester 5 kg. 1 o tolym l ymhdone 32g 8 wherein R ishydrogen, lower alkyl or lower alkoxy,

Talc 30 R is lower alkyl, phenyl or phenyl-lower alkyl, X is Magnesiumstearate 80 gm. hydrogen, hydroxy, lower alkanoyloxy, benzoyloxy orphenyl-lower alkanoyloxy, and n is 0 or I. 5 2. A compound as in claim 1wherein R and X each is hydrogen and n is 0.

3. A compound as in claim 1 wherein R is t-butylr The active substanceis mixed with the lactose and granulated with an ethanol solution of thepolyvinyl pyrrolidone. The wet material is screened, then dried at 4 Acnmpound as in claim 2 wherein 1 is LbuwL 45. The dried material isscreened and admixed with 5 A compound as i l im 1 wherein R is hydrogenthe talc and magnesium stearate. The mixture is com- X i lowera|kan0y|0xy and n is (L pressed in a tabletting machine to obtain 10,000tablets 6. A compound as in claim 5 wherein R is t-butyl. each weighinga total of 630 mg. and containing 500 7. A compound as in claim 5wherein the lower almg. of active ingredient. kanoyloxy group is acetoxyand R is t-butyl.

What is claimed is: I! t t

2. A compound as in claim 1 wherein R and X each is hydrogen and n is 0.3. A compound as in claim 1 wherein R1 is t-butyl.
 4. A compound as inclaim 2 wherein R1 is t-butyl.
 5. A compound as in claim 1 wherein R ishydrogen, X is lower alkanoyloxy and n is
 0. 6. A compound as in claim 5wherein R1 is t-butyl.
 7. A compound as in claim 5 wherein the loweralkanoyloxy group is acetoxy and R1 is t-butyl.